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Mycobacterial biofilm is a structured community of bacterial cells enclosed in a self-produced polymeric matrix and adherent to an inert or living surface, which constitutes a protected mode of growth that allows survival in hostile environment. Biofilms can be defined as communities of mycobacteria attached to a surface. It is clear that microorganisms undergo profound changes during their transition from planktonic (free-swimming) organisms to cells that are part of a complex, surface-attached community. These changes are reflected in the new phenotypic characteristics developed by biofilm mycobacteria and occur in response to a variety of environmental signals. The biofilm-forming microorganisms have been shown to elicit specific mechanisms for initial attachment to a surface, formation of micro colony leading to development of three-dimensional structure of mature biofilm. They differ from their free-living counterparts in their growth rate, composition and increased resistance to biocides, antibiotics and antibodies by virtue of up regulation and/or down regulation of approximately 40% of their genes. This makes them highly difficult to eradicate with therapeutic doses of antimicrobial agents. A greater understanding of mechanism of their formation and survival under sessile environments may help in devising control strategies.